A major quest in health care today is elucidating the mechanism for Autism Spectrum Disorders (ASD), one of the leading causes of autistic behavior occurring in 1:150 births. Rett Syndrome (RTT) is a developmental disorder of the brain that occurs in females and is responsible for mental retardation and autistic behavior. RTT falls under the umbrella of ASD. Mutations in the X-linked MeCP2 protein situated on the X chromosome are the cause of this genetic disorder. Phenotypic variation ranging from mild to severe manifestations is observed in RTT. A major determinant of this clinical variability is the pattern of X-chromosome inactivation (XCI), a crucial epigenetic process that occurs early in development to balance the gene dose between XX females and XY males. Female cells undergo a chromosomal-wide silencing of one X chromosome at random to equalize the dose. Favorable XCI that silences the X chromosome with the mutant MeCP2 gene will lessen the severity of symptoms in girls with RTT. Insight into how XCI is regulated will provide understanding in the pathogenesis of RTT the may offer hope in the amelioration of severe phenotypes. These studies have profound impact on allelic expression diseases such as genomic imprinted gene disorders, such as Angelman<s and Prader-Willi Syndromes, and X-linked mental retardation. We have discovered choice trans-factors that skew XCI in the mouse. We will use the powerful system of mouse embryonic stem (ES) cells to investigate how XCI might skew favorably to decrease the expression of the genetically inherited mutant MeCP2 gene. The role of the chromatin insulator Ctcf and its protein partners will be investigated in the transcriptional regulation of MeCP2. Prior to a choice of X chromosome to be inactivated, the two female X chromosomes transiently touch to set a mutually exclusive designation of active versus inactive X chromosome. This X-X kissing is correlated with ES cell differentiation and our studies show it is mediated by a pluripotent factor that is a co-factor of Ctcf. We will examine this process in normal and mutant MeCP2 female ES cells. We will attempt to manipulate X choice by altering the expression of the trans-factors crucial for this process. This grant synergizes the fields of medicine, genetics and epigenetics to ask a translational question: how can we lessen the severity of a genetic disease? Our long-term goal is to ask experimentally how one of the two female X chromosome is chosen to be inactivated in normal development and armed with this knowledge, we can alter the fate of inactivation to the mutated X chromosome. Can we selectively activate the wildtype MeCP2 gene using chromatin insulation? Knowledge these basic mechanisms will further our understanding of Autism Spectrum disorders that are so prevalent today.